Program in Macromolecular Structure, Motion, Control

高分子结构、运动、控制程序

• 批准号: 7297748
• 负责人: SCOTT A STROBEL
• 金额: $ 19万
• 依托单位: YALE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2006
• 资助国家: 美国
• 起止时间: 2006-04-25 至 2009-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7297748
• 关键词: RNA; RNA splicing; X ray crystallography; active sites; chemical structure; conformation; gene rearrangement; macromolecule; metalloenzyme; molecular dynamics; ribonucleoproteins; small molecule

RNA splicing is the removal of an intron and the simultaneous ligation of its flanking exons in the generation of mature cellular RNA molecules. Splicing provides a critical level of genetic control. Through alternative splicing, the proteome of a higher eukaryote is substantially more complex than the number of genes in its genome. The importance of RNA splicing to human health is manifest by the observation that at least 15% of point mutations leading to heritable human diseases cause defects in pre-mRNA splicing. While most introns are removed by the spliceosome, some introns are able to catalyze their own removal from the primary transcript. The discovery of the group I class of introns provided the first indication that not all enzymes are proteins. Their existence proves that RNA can select 5' and 3' splice sites and catalyze the two transesterification reactions of intron removal. Our understanding of the structural basis of RNA splicing is still in its infancy, but in 2004, almost 25 years after its initial discovery, the first crystal structure of a complete group I intron in complex with both its 5' and 3' exons was finally determined. This result will now be exploited to achieve several important objectives relevant to the structural basis of RNA splicing and the dynamic conformational rearrangements that occur during the splicing process. The overriding goals of these studies are: (i) to determine the X-ray crystal structure of each step in an RNA splicing pathway, (ii) to explain how RNA tertiary structure is formed and active sites created in the absence of proteins, (iii) to reveal how metal ions contribute to RNA catalysis and how alteration of ligands affects metal ion specificity, (iv) to visualize the nature of the transition state of the phosphoryl transfer reaction promoted during the 5'-exon cleavage and exon ligation reactions, and (v) to explore how group I intron splicing is facilitated by protein cofactors. Many ribonucleoprotein complexes are expected to undergo complex conformational changes during their function, so understanding how an RNA is reconfigured during a multi-step splicing reaction will provide a valuable precedent for considering these complex dynamic processes. RNA enzymes, or ribozymes, are the molecules most likely to be the progenitors of modern biological catalysts, and understanding how they promote their reactions will provide critical insight into enzymological function. This series of structural snapshots will reveal principles of RNA folding, structural dynamics, and metal mediated catalysis, principles that are certain to have parallels in most cellular machines that include RNA.

RNA剪接是去除内含子并同时连接其两侧外显子的过程 成熟的细胞RNA分子。剪接提供了一个关键的遗传控制水平。通过可选的剪接， 高等真核生物的蛋白质组比其基因组中的基因数量要复杂得多。这个 观察到至少15%的点突变对人类健康的重要性是显而易见的 导致人类遗传性疾病会导致前mRNA剪接缺陷。虽然大多数内含子是由 剪接体，一些内含子能够催化自己从初级转录本上移除。人类的发现 第一类内含子首次表明并非所有的酶都是蛋白质。它们的存在证明了 RNA可以选择5‘和3’剪接位点，催化内含子去除的两个酯交换反应。我们的 对RNA剪接的结构基础的了解仍处于初级阶段，但在2004年，在其 初步发现，一个完整的I族内含子及其5‘和3’外显子的复合体的第一个晶体结构是 终于确定了。现在将利用这一结果来实现与 RNA剪接的结构基础和剪接过程中的动态构象重排 进程。这些研究的首要目标是：(I)确定RNA中每一步的X射线晶体结构 剪接途径，(Ii)解释在没有剪接的情况下，RNA三级结构是如何形成的，活性位点是如何产生的 蛋白质，(Iii)揭示金属离子如何对RNA催化起作用以及配体的改变如何影响金属离子 专一性，(Iv)直观地显示在 5‘-外显子切割和外显子连接反应，以及(V)探讨蛋白质如何促进I组内含子剪接 辅因。许多核糖核蛋白复合体在其形成过程中预计会经历复杂的构象变化 功能，因此了解RNA在多步骤剪接反应中是如何重新配置的将提供有价值的 这是考虑这些复杂的动态过程的先例。核糖核酸酶或核酶是最重要的分子。 可能是现代生物催化剂的先驱，了解它们如何促进他们的反应将 提供对酶的功能的关键洞察。这一系列的结构快照将揭示 RNA折叠、结构动力学和金属催化，这些原理在大多数情况下肯定是相似的 包括RNA在内的细胞机器。